Direct visualization of membrane leakage induced by the antibiotic peptides: Maculatin, citropin, and aurein
Direct visualization of membrane leakage induced by the antibiotic peptides: Maculatin, citropin, and aurein
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DOI：
10.1529/biophysj.105.066589
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发表时间：
2005-09-01
影响因子：
3.4
通讯作者：
Bagatolli, LA
中科院分区：
文献类型：
--
作者：
Ambroggio, EE;Separovic, F;Bagatolli, LA
抗生素肽引起的膜裂解通常通过两种不同的模型来合理化：所谓的地毯模型和成孔模型。我们在此报道了澳大利亚树蛙抗生素肽Maculatin 1.1, Citropin 1.1和Aurein 1.2在POPC或POPC/POPG模型膜上的裂解活性。荧光光谱渗漏实验表明，无论脂膜组成如何，Maculatin诱导50%探针渗漏所需的肽/脂质mol比都小于aurin或Citropin。为了进一步了解这些肽的裂解机制，我们使用共聚焦荧光显微镜进行了单囊泡实验。在这些实验中，我们观察了不同分子量（水溶性）的荧光标记物在肽暴露后渗漏的时间过程。我们得出结论，Maculatin及其相关肽显示出一种成孔机制（与高分子量标记物相比，小荧光探针的差异泄漏）。相反，Citropin和aurin引起囊泡破裂的完全膜不稳定，而没有连续的探针泄漏，这种效应可以被分配到溶解作用的地毯机制。此外，为了研究脯氨酸残基与Maculatin膜作用特性的相关性，我们对Maculatin-Ala和Maculatin-Gly采用了相同的实验方法（将Pro-15分别替换为Ala或Gly）。虽然类似的肽/脂质摩尔比可以诱导POPC膜50%的渗漏，但与自然发生的Maculatin序列相比，Maculatin- ala和Maculatin- gly在POPC/POPG （1:1 mol）膜中的裂解活性降低。在Maculatin中观察到，Maculatin- ala和Maculatin- gly的裂解作用与膜上孔状结构的形成保持一致，独立于脂质组成。
Membrane lysis caused by antibiotic peptides is often rationalized by means of two different models: the so-called carpet model and the pore-forming model. We report here on the lytic activity of antibiotic peptides from Australian tree frogs, Maculatin 1.1, Citropin 1.1, and Aurein 1.2, on POPC or POPC/POPG model membranes. Leakage experiments using fluorescence spectroscopy indicated that the peptide/lipid mol ratio necessary to induce 50% of probe leakage was smaller for Maculatin compared with Aurein or Citropin, regardless of lipid membrane composition. To gain further insight into the lytic mechanism of these peptides we performed single vesicle experiments using confocal fluorescence microscopy. In these experiments, the time course of leakage for different molecular weight ( water soluble) fluorescent markers incorporated inside of single giant unilamellar vesicles is observed after peptide exposure. We conclude that Maculatin and its related peptides demonstrate a pore-forming mechanism ( differential leakage of small fluorescent probe compared with high molecular weight markers). Conversely, Citropin and Aurein provoke a total membrane destabilization with vesicle burst without sequential probe leakage, an effect that can be assigned to a carpeting mechanism of lytic action. Additionally, to study the relevance of the proline residue on the membrane-action properties of Maculatin, the same experimental approach was used for Maculatin-Ala and Maculatin-Gly (Pro-15 was replaced by Ala or Gly, respectively). Although a similar peptide/lipid mol ratio was necessary to induce 50% of leakage for POPC membranes, the lytic activity of Maculatin-Ala and Maculatin-Gly decreased in POPC/POPG ( 1: 1 mol) membranes compared with that observed for the naturally occurring Maculatin sequence. As observed for Maculatin, the lytic action of Maculatin-Ala and Maculatin-Gly is in keeping with the formation of pore-like structures at the membrane independently of lipid composition.